反义寡核苷酸抑制survivin基因表达及肝癌细胞生长的研究

The Effect of Antisense Oligonucleotide on the Expression of Survivin Gene and Proliferation of Human Hepatocellular Carcinoma Cell Line SMMC-7721

目的:采用反义寡核苷酸抑制肝癌细胞中survivin基因的表达,研究其对肝癌细胞生长的作用。方法:采用脂质 体介导survivin反义寡核苷酸转染人肝癌细胞株SMMC-7721细胞,Western blot及原位杂交方法检测survivin蛋白及mRNA表 达,流式细胞仪检测凋亡细胞比率,检测转染前后细胞贴壁率变化,并绘制细胞生长曲线。结果:survivin反义寡核苷酸转染 后,肝癌细胞survivin蛋白及mRNA表达均显著降低,细胞贴壁率显著降低,细胞增殖活性明显受抑,细胞凋亡率显著增加。 结论:survivin反义寡核苷酸转染可以有效降低细胞内survivin基因的表达,诱导细胞发生凋亡,抑制肝癌细胞生长。

Objective: To investigate the inhibitory effect of survivin antisense oligonucleotide (ASODN) on the expression of survivin gene and proliferation of human hepatocellular carcinoma cell line SMMC-7721. Methods: The 20 mer antisense oligonucleotide (ASODN) targeted to the promoter region of survivin mRNA was designed and synthesized. The expression of survivin gene in hepatocellular carcinoma cell line SMMC-7721 was blocked by means of ASODN transfection mediated by DOTAP liposomal reagent. The changes of survivin protein and mRNA expression after transfection were as-sessd by Western blot and in situ hybridization, respectively. The apoptotic rate was detected by flow cytometer. The changes of cell adherent rate, cell growth activity, and the inhibitory rate of cell growth were also studied. Results: The expression of survivin protein and mRNA were decreased markedly after survivin ASODN transfection. Meanwhile, the cell adherent rate also decreased markedly while the apoptotic rate increased markedly. Conclusions: Transfection of ASODN targeted to the promotor region of survivin mRNA by DOTAP liposomal transfection reagent could down-regulated the expression of survivin protein and mRNA significantly in 7721 cell line and inhibit the proliferation of cancer cells. Survivin could be an important target in the therapy of hepatocellular carcinoma.